In the United States, drug addiction is a major medical problem and has significant implications for public health and society with over one million people addicted to heroin. Physiological systems are disrupted by drugs of abuse, and these disruptions contribute to drug addiction and relapse. Epidemiological studies have indicated that non-genetic factors contribute 50-80% of the risk of developing addiction. Among these non- genetic factors are epigenetic factors which include methylation of cytosines that reside in cytosine:guanine (CpG) dinucleotides in genomic DNA. This common epigenetic mechanism is known to repress gene expression. Methylated DNA recruits methyl-CpG repressor protein and enzymes responsible for altering histones, both repressing gene expression probably through the inhibition of transcription factor binding. Furthermore, methylation is important in the expression of imprinted genes and in the development of some cancers. Although methylation patterns are mostly stable after birth, exogenous factors, such as dietary factors, can alter DNA methylation state and transcriptional activity. In addition, the chromatin structure of several promoters has been shown to be altered by cocaine (dependent on the treatment regime). We hypothesize that DNA methylation patterns in the promoter regions of genes known to be involved in the effects of drugs of abuse, or associated with predisposition to drug abuse, will differ between former opiate addicts (without or without cocaine addiction) and controls. This difference in the methylation patterns may be due to the drug addiction itself or due to a predisposition to drug abuse. In this application we propose to 1) study the DNA methylation patterns in former opiate addicts (without or with cocaine addiction) and control subjects and 2) if a methylation state difference is found, analyze the expression of these genes in lymphocytes from former severe opiate addicts and controls. The identification of a methylation state difference and a relation of this methylation pattern to expression between addicted patients and controls would be a provocative and exciting finding that would lead to future studies. An understanding of this epigenetic mechanism in heroin addiction is essential and may lead to the elucidation of novel pharmacotherapies for the treatment of opiate addiction. Findings of the studies in this application may help to alleviate individual suffering and the tremendous cost to society caused by drug and alcohol addiction. [unreadable] [unreadable] [unreadable]